The long term objective of this research proposal is to gin new insights into the detailed molecular and cellular events which are triggered by recognition of foreign antigens and which generate the appropriate immune responses. In particular, the studies will be aimed at understanding the molecular and cellular mechanisms of specific cell:cell interactions, and how such interactions cause the activation and differentiation of Beta cells by T-helper (Th) cells, as well as T cell selection in the thymus. These studies will be conducted primarily at the single cell level using double and triple immunofluorescence microscopy. Such cell biological techniques already demonstrated the in vitro formation of specific stable Th:Beta-cell conjugates, and have identified several important early intracellular and membrane events triggered by such T:B interactions. The results of these studies will increase our knowledge of the mechanisms by which cells of the immune system can protect against invading microorganisms, viral infections, and some malignancies. The specific aims of the proposed research are: 1. To study intracellular and membrane rearrangements induced by the interactions of cloned Th (Th1, Th2) cells with resting Beta cells which will be either Ag/MHC specific, non-specific or bystander (non-specific in the presence of specific) Beta cells. 2. To study the spatial and temporal induction of lymphokine (e.g. IL2, IL4) secretion in specific and mixed (bystander) T:B conjugates. 3. To study the activation and differentiation of resting Beta cells in T;Beta conjugates, including complex conjugates in which several Beta cells are simultaneously bound to one T cell. 4. To continue to study the signalling mechanisms that trigger the intracellular and membrane rearrangements, which were identified before in specific T: Beta and CTL:TC conjugates. 5. To study the signalling mechanisms in defined populations of immature T cells.